Olfactory neurons use a large repertoire of G protein-coupled odorant receptors to detect thousands of odors. Ligand-stimulated receptor phosphorylation is a major regulatory mechanism to receptor function for many G-protein coupled receptors (GPCRs) and is essential to the termination of GPCR signal transduction. As with other G protein-coupled signaling, olfactory transduction may use the receptor phosphorylation mechanism to regulate the response to odor stimuli. Yet, there is still lack of a direct and unambiguous evidence for odorant-stimulated odorant receptor phosphorylation. How and how much odorant-stimulated odorant receptor phosphorylation contributes to olfactory signal transduction remains largely unknown. The overall goal of this project is to understand how olfactory signal transduction is regulated at the level of the odorant receptor. Two specific aims proposed in this project include: 1) to demonstrate odorant-stimulated odorant receptor phosphorylation; and 2) to reveal the role of odorant-stimulated odorant receptor phosphorylation in olfactory signal transduction. The first aim will be achieved by expressing an epitope tagged odorant receptor with known ligand specificity in all olfactory neurons in mice through a gene targeting approach. The odorant-stimulated odorant receptor phosphorylation will be displayed by detecting the phosphorylated receptor protein using well-defined antibodies against the epitope tag. The second aim will be achieved by expressing a mutated odorant receptor, in which the potential phosphorylation residues are mutated, in all olfactory neurons in mice. The odor-evoked responses in these mice will be measured by electrophysiological recordings and be compared to responses in the mouse carrying ubiquitous expression of the wild-type receptor. The proposed research will enhance our understanding of how olfactory neurons control their sensitivity and response dynamics to odor stimuli.